An investigation of the role of metabolism in dapsone-induced methaemoglobinaemia using a two compartment in vitro test system.
1. We have utilized a two compartment system in which two teflon chambers are separated by a semi-permeable membrane in order to investigate the role of metabolism in dapsone-induced methaemoglobinaemia. Compartment A contained a drug metabolizing system (microsomes prepared from human liver +/- NADPH), whilst compartment B contained target cells (human red cells). 2. Incubation of dapsone (1-100 microM) with human liver microsomes (2 mg protein) and NADPH (1 mM) in compartment A (final volume 500 microliters) led to a concentration-dependent increase in the methaemoglobinaemia (15.4-18.9% at 100 microM) compared with control (2.3 +/- 0.4%) detected in the red cells within compartment B. In the absence of NADPH dapsone had no effect. 3. Of the putative dapsone metabolites investigated, only dapsone-hydroxylamine caused methaemoglobin formation in the absence of NADPH (40.6 +/- 6.3% with 100 microM). However, methaemoglobin was also detected when monoacetyl-dapsone, 4-amino-4'-nitro-diphenylsulphone and 4-aminoacetyl-4'-nitro-diphenylsulphone were incubated with human liver microsomes in the presence of NADPH. 4 Dapsone-dependent methaemoglobin formation was inhibited by addition of ketoconazole (1-1000 microM) to compartment A, with IC50 values of 285 and 806 microM for the two liver microsomal samples studied. In contrast, methaemoglobin formation was not inhibited by cimetidine or a number of drugs pharmacologically-related to dapsone. The presence of glutathione or ascorbate (500 microM) did not alter the level of methaemoglobin observed.
Tingle
M D
MD
Department of Pharmacology and Therapeutics, University of Liverpool.
Coleman
M D
MD
Park
B K
BK
D002927:Cimetidine / Q000494:pharmacology; D003622:Dapsone / Q000378:metabolism* / Q000633:toxicity; D004790:Enzyme Induction / Q000187:drug effects; D006801:Humans; D007654:Ketoconazole / Q000494:pharmacology; D008706:Methemoglobin / Q000096:biosynthesis*; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009249:NADP / Q000378:metabolism
D009249:NADP; D002927:Cimetidine; D003622:Dapsone; D008706:Methemoglobin; D007654:Ketoconazole